Rat peritoneal mast cells incubated in Hanks medium for various periods released histamine into the medium. This release was higher in immature mast cells than mature ones. After incubation of mast cells with 14C(ring) and 14C (carboxyl)-histidine, the amount of unlabeled histamine retained by the cells was much less than the 14CO2 released. Both the appearance of unlabeled histamine in the medium and the synthesis of labeled histamine were reduced by Alpha-fluoromethyl histidine and NSD-1055. From these studies we conclude that a significant portion of the newly synthesized histamine in mast cells is released into the extracellular medium. Histidine decarboxylase activity from different tissues of rat was inhibited by increasing concentrations of Alpha-fluoromethyl histidine. Prolonged incubation of the enzymes with the inhibitor enhanced the inhibitory action of low concentrations of Alpha-fluoromethyl histidine and so did the presence of pyridoxal phosphate during incubation. Studies with elutriated rat basophilic leukemia (2H3) cells showed that these cells have low histamine content but the histamine synthetic activity is high. The pattern of histamine content and histamine synthetic activity are similar in the elutriated fractions.